As will be discussed hereinafter, the present inventors have made several proposals about apparatuses for detecting chemotaxis of cells by which chemotaxis of cells can be detected with the use of a microquantity of cells by supplying a cell suspension into a well, supplying a specimen into another well and examining whether or not the cells migrate toward the well containing the specimen via a channel provided between the wells. Use of an apparatus for detecting chemotaxis of cells enabling the observation and detection at the single cell level and allowing measurement with the use of about 10 to 100 cells, if possible, offers such advantages that scarce cells can be easily examined and cell reactions can be quantitatively analyzed and studied. In a structure wherein individual wells, which are connected to each other via channels, have each a cell injection port and a specimen injection port, wells together form a connecting tube via the channels and, in its turn, a liquid frequently migrates in the channels. In the step of injecting a specimen such as cells into a well, namely, there frequently arises an increase in pressure, which results in unexpected migration of cells or the specimen in many cases. In the case wherein wells are not completely leveled after the injection or micromovements occur on the liquid level due to, for example, vibration, the transportation of the liquid in the channel is amplified so as to frequently cause migration of cells or a specimen. Such unexpected migration of cells or a specimen causes confusion in the judgment whether the specimen is a chemotactic factor or not. To accurately detect the migration of cells, which perceive the concentration gradient due to the diffusion of the specimen, toward a well containing a specimen, it is therefore required to strictly prevent the unexpected migration of the liquid in the wells involving the channels. To further exactly understand the chemotaxis of cells, it is desirable that individual cells are located in the same state in a well, i.e., forming a line toward a channel, at starting.
The present inventors have previously proposed that, in the case of injecting or aspirating a sample into a well having a tube for injecting/aspirating a sample such as a cell or a chemotactic factor with a micropipette or the like, a rapid change in pressure in a well can be relieved and unexpected migration of the sample in the well can be prevented by providing another tube connecting to the tube for injecting or aspirating the sample (JP-A-2002-159287). In this case, use is made of the structure wherein the pressure in injecting or aspirating a sample is dispersed through the connecting tube.
The present inventors have further proposed a microsample treatment apparatus such as an apparatus for detecting chemotaxis of cells wherein, in a structure comprising a plural number of wells being connected to each other via a part having resistance to fluids and individual wells being provided with tubes for injecting/aspirating a sample and, if necessary, tubes for relieving pressure changes at the injection/aspiration, these tubes have a space in common at the top ends thereof in which a liquid can be held (WO 02/46356). By employing the structure wherein all of the tubes provided in individual wells have a space in common at the top ends thereof in which a liquid can be held, the migration of a liquid at the step of injecting a sample and thereafter can be prevented. In this apparatus, moreover, the positions of the injected cells in a well can be controlled so as to align the cells along the start line at one end of a channel. For this purpose, this apparatus has a means of precisely controlling the injection/discharge of a liquid in a well.
To align cells injected into a well along the start line at one end of a channel, the liquid in the well should be moved and delicate control is required therefor. To prevent the subsequent migration of the liquid in the channel, it is also required to return the liquid into the space at the top ends of the tubes. That is, complicated and strictly controlled procedures are necessary therefor.
In addition to the present inventors' proposals as described above, there has been known an apparatus for detecting chemotaxis of cells wherein wells containing cells and a specimen are connected to each other via channels and the wells are provided with a means of sealing injection ports for the cells and the specimen if necessary (U.S. Pat. No. 5,744,366). In this apparatus, however, cell positioning cannot be controlled in a well and it is therefore impossible to adjust the conditions of the cells at starting.
An object of the present invention is to further improve such an apparatus and thus provide a structure for detecting chemotaxis of cells at an elevated accuracy with the use of a microquantity of cells. Namely, an object of the present invention is to provide an apparatus for detecting chemotaxis of cells by which cells or a specimen can be easily injected and position control of the injected cells can be easily carried out while ensuring the prevention of unexpected migration of the cells definitely positioned in a well or the injected specimen so that a stable concentration gradient due to the diffusion of the specimen can be maintained and which ensures further automated operation and controlling.